Browsing by Author "Huang, Hsien-Da"

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A computational approach for identifying microRNA-target interactions using high-throughput CLIP and PAR-CLIP sequencing
(2013-01-21)
Chou, Chih-Hung
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Lin, Feng-Mao
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Chou, Min-Te
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Hsu, Sheng-Da
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Chang, Tzu-Hao
Background: MicroRNAs (miRNAs) play a critical role in down-regulating gene expression. By coupling with Argonaute family proteins, miRNAs bind to target sites on mRNAs and employ translational repression. A large amount of miRNA-target interactions (MTIs) have been identified by the crosslinking and immunoprecipitation (CLIP) and the photoactivatable-ribonucleoside-enhanced CLIP (PAR-CLIP) along with the next-generation sequencing (NGS). PAR-CLIP shows high efficiency of RNA co-immunoprecipitation, but it also lead to T to C conversion in miRNA-RNA-protein crosslinking regions. This artificial error obviously reduces the mappability of reads. However, a specific tool to analyze CLIP and PAR-CLIP data that takes T to C conversion into account is still in need. Results: We herein propose the first CLIP and PAR-CLIP sequencing analysis platform specifically for miRNA target analysis, namely miRTarCLIP. From scratch, it automatically removes adaptor sequences from raw reads, filters low quality reads, reverts C to T, aligns reads to 3'UTRs, scans for read clusters, identifies high confidence miRNA target sites, and provides annotations from external databases. With multi-threading techniques and our novel C to T reversion procedure, miRTarCLIP greatly reduces the running time comparing to conventional approaches. In addition, miRTarCLIP serves with a web-based interface to provide better user experiences in browsing and searching targets of interested miRNAs. To demonstrate the superior functionality of miRTarCLIP, we applied miRTarCLIP to two public available CLIP and PAR-CLIP sequencing datasets. miRTarCLIP not only shows comparable results to that of other existing tools in a much faster speed, but also reveals interesting features among these putative target sites. Specifically, we used miRTarCLIP to disclose that T to C conversion within position 1-7 and that within position 8-14 of miRNA target sites are significantly different (p value = 0.02), and even more significant when focusing on sites targeted by top 102 highly expressed miRNAs only (p value = 0.01). These results comply with previous findings and further suggest that combining miRNA expression and PAR-CLIP data can improve accuracy of the miRNA target prediction. Conclusion: To sum up, we devised a systematic approach for mining miRNA-target sites from CLIP-seq and PAR-CLIP sequencing data, and integrated the workflow with a graphical web-based browser, which provides a user friendly interface and detailed annotations of MTIs. We also showed through real-life examples that miRTarCLIP is a powerful tool for understanding miRNAs. Our integrated tool can be accessed online freely at http://miRTarCLIP.mbc.nctu.edu.tw.
A Human DNA methylation site predictor base on SVM
(2009)
Sun, Yi-Ming
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Liu, Baw-Jhiune
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Liao, Wei-Li
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Chang, Cheng-Wei
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Huang, Hsien-Da
During gene expression, transcription factors are unable to bind to a transcription binding site (TFBS) involved in regulation if DNA methylation has occurred at the TFBS. Methyl-CpG-binding proteins may also occupy the TFBS and prevent the functioning of a transcription factor. Thus, the methylation status of CpG sites is an important issue when trying to understand gene regulation and shows strong correlation with the TFBS involved. In addition, CpG islands would seem to undergo cell-specific and tissue-specific methylation. Such differential methylation is presented at numerous genetic loci that are essential for development. Current DNA methylation site prediction tools need to be improved so that they include TFBS features and have greater accuracy in terms of the DNA region that is involved in methylation. We developed models that compare the differences across these regions and tissues. The TFBSs, DNA properties and DNA distribution were used as features for this classification. From the results, we found some TFBSs that were able to discriminate whether a sequence was methylated or not. The sensitivity, specificity and accuracy estimated using 10-fold cross validation were 90.8%, 80.54%, and 86.07%, respectively. Thus, for these four regions and twelve tissues, the performance levels (ACC) were all greater than 80%. We propose that the differential features or methylations vary between the different regions because the features common to each DNA region made up only 50% of the top 70 features. An online predictor based on EpiMeP is available at http://140.115.51.41/EpiMeP/. Supplementary file is available at http://140.115.51.41/EpiMeP/supplementary.doc.
A new method for post Genome-Wide Association Study (GWAS) analysis of colorectal cancer in Taiwan
(2013-04-10)
Wang, Hwei-Ming
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Chang, Tzu-Hao
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Lin, Feng-Mao
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Chao, Te-Hsin
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Huang, Wei-Chih
Recently, single nucleotide polymorphisms (SNPs) located in specific loci or genes have been identified associated with susceptibility to colorectal cancer (CRC) in Genome-Wide Association Studies (GWAS). However, in different ethnicities and regions, the genetic variations and the environmental factors can widely vary. Therefore, here we propose a post-GWAS analysis method to investigate the CRC susceptibility SNPs in Taiwan by conducting a replication analysis and bioinformatics analysis. One hundred and forty-four significant SNPs from published GWAS results were collected by a literature survey, and two hundred and eighteen CRC samples and 385 normal samples were collected for post-GWAS analysis. Finally, twenty-six significant SNPs were identified and reported as associated with susceptibility to colorectal cancer, other cancers, obesity, and celiac disease in a previous GWAS study. Functional analysis results of 26 SNPs indicate that most biological processes identified are involved in regulating immune responses and apoptosis. In addition, an efficient prediction model was constructed by applying Jackknife feature selection and ANOVA testing. As compared to another risk prediction model of CRC for European Caucasians population, which performs 0.616 of AUC by using 54 SNPs, the proposed model shows good performance in predicting CRC risk within the Taiwanese population, i.e., 0.724 AUC by using 16 SNPs. We believe that the proposed risk prediction model is highly promising for predicting CRC risk within the Taiwanese population. In addition, the functional analysis results could be helpful to explore the potential associated regulatory mechanisms that may be involved in CRC development. (c) 2012 Elsevier B.V. All rights reserved.
A novel role of CPEB3 in regulating EGFR gene transcription via association with Stat5b in neurons
(2010-11-01)
Peng, Shu-Chun
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Lai, Yen-Ting
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Huang, Hsi-Yuan
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Huang, Hsien-Da
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Huang, Yu-Shuian
CPEB3 is a sequence-specific RNA-binding protein and represses translation of its target mRNAs in neurons. Here, we have identified a novel function of CPEB3 as to interact with Stat5b and inhibit its transcription activity in the nucleus without disrupting dimerization, DNA binding and nuclear localization of Stat5b. Moreover, CPEB3 is a nucleocytoplasm-shuttling protein with predominant residence in the cytoplasm; whereas activation of NMDA receptors accumulates CPEB3 in the nucleus. Using the knockdown approach, we have found the receptor tyrosine kinase, EGFR, is a target gene transcriptionally activated by Stat5b and downregulated by CPEB3 in neurons. The increased EGFR expression in CPEB3 knockdown neurons, when stimulated with EGF, alters the kinetics of downstream signaling. Taken together, CPEB3 has a novel function in the nucleus as to suppress Stat5b-dependent EGFR gene transcription. Consequently, EGFR signaling is negatively regulated by CPEB3 in neurons.
A novel role of CPEB3 in regulating EGFR gene transcription via association with Stat5b in neurons
(2010-11-01)
Peng, Shu-Chun
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Lai, Yen-Ting
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Huang, Hsi-Yuan
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Huang, Hsien-Da
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Huang, Yu-Shuian
CPEB3 is a sequence-specific RNA-binding protein and represses translation of its target mRNAs in neurons. Here, we have identified a novel function of CPEB3 as to interact with Stat5b and inhibit its transcription activity in the nucleus without disrupting dimerization, DNA binding and nuclear localization of Stat5b. Moreover, CPEB3 is a nucleocytoplasm-shuttling protein with predominant residence in the cytoplasm; whereas activation of NMDA receptors accumulates CPEB3 in the nucleus. Using the knockdown approach, we have found the receptor tyrosine kinase, EGFR, is a target gene transcriptionally activated by Stat5b and downregulated by CPEB3 in neurons. The increased EGFR expression in CPEB3 knockdown neurons, when stimulated with EGF, alters the kinetics of downstream signaling. Taken together, CPEB3 has a novel function in the nucleus as to suppress Stat5b-dependent EGFR gene transcription. Consequently, EGFR signaling is negatively regulated by CPEB3 in neurons.
A panel of eight microRNAs is a good predictive parameter for triple-negative breast cancer relapse
(2020-01-01)
Hong, Hsiao-Chin
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Chuang, Cheng-Hsun
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Huang, Wei-Chih
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Weng, Shun-Long
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Chen, Chia-Hung
Rationale: Triple-negative breast cancer (TNBC), which has the highest recurrence rate and shortest survival time of all breast cancers, is in urgent need of a risk assessment method to determine an accurate treatment course. Recently, miRNA expression patterns have been identified as potential biomarkers for diagnosis, prognosis, and personalized therapy. Here, we investigate a combination of candidate miRNAs as a clinically applicable signature that can precisely predict relapse in TNBC patients after surgery. Methods: Four total cohorts of training (TCGA_TNBC and GEOD-40525) and validation (GSE40049 and GSE19783) datasets were analyzed with logistic regression and Gaussian mixture analyses. We established a miRNA signature risk model and identified an 8-miRNA signature for the prediction of TNBC relapse. Results: The miRNA signature risk model identified ten candidate miRNAs in the training set. By combining 8 of the 10 miRNAs (miR-139-5p, miR-10b-5p, miR-486-5p, miR-455-3p, miR-107, miR-146b-5p, miR-324-5p and miR-20a-5p), an accurate predictive model of relapse in TNBC patients was established and was highly correlated with prognosis (AUC of 0.80). Subsequently, this 8-miRNA signature prognosticated relapse in the two validation sets with AUCs of 0.89 and 0.90. Conclusion: The 8-miRNA signature predictive model may help clinicians provide a prognosis for TNBC patients with a high risk of recurrence after surgery and provide further personalized treatment to decrease the chance of relapse.
A systematic review of microRNA expression profiling studies in human gastric cancer
(2014-08-01)
Shrestha, Sirjana
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Hsu, Sheng-Da
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Huang, Wei-Yun
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Huang, Hsi-Yuan
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Chen, WenLiang
Gastric cancer (GC) is the second leading cause of global cancer mortality. Most GC patients are diagnosed with advanced-stage disease and show extremely poor prognosis. All of the GC research has a common interest to search for the specific and sensitive biomarkers for early diagnosis of GC. Number of microRNAs play important role in GC. We carried out a systematic review of published miRNA profiling studies that compared the miRNA expression profiles between GC tissues and paired noncancerous gastric tissue. A vote-counting strategy was followed with the collection of information like total number of studies reporting differential expression of miRNA, total number of tissue samples used in the studies, direction of differential expression and fold change. A total of 352 differentially expressed microRNAs were reported in the 14 microRNA expression profiling studies that compared GC tissues with normal tissues with 120 microRNAs reported at least in two studies. In the group of consistently reported microRNAs, miR-21 was reported upregulated in 10 studies followed by miR-25, miR-92, and miR-223 upregulated in eight studies. MiR-375 and miR-148a were found downregulated in six and five studies, respectively, followed by miR-638 in four studies. MiR-107 and miR-103 were reported in nine and eight studies, respectively, but their expression were inconsistent. From this study, the most consistently reported upregulated microRNA was found to be miR-21. This systematic review study of human GC microRNA expression profiling studies would provide information on microRNAs with potential role as the biomarkers in gastric cancer.
A Three-MicroRNA Signature as a Potential Biomarker for the Early Detection of Oral Cancer
(2018-03-01)
Chang, Yi-An
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Weng, Shun-Long
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Yang, Shun-Fa
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Chou, Chih-Hung
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Huang, Wei-Chih
Oral squamous cell carcinoma (OSCC) is often diagnosed at a late stage and may be malignantly transformed from oral leukoplakia (OL). This study aimed to identify potential plasma microRNAs (miRNAs) for the early detection of oral cancer. Plasma from normal, OL, and OSCC patients were evaluated. Small RNA sequencing was used to screen the differently expressed miRNAs among the groups. Next, these miRNAs were validated with individual samples by quantitative real-time polymerase chain reaction (qRT-PCR) assays in the training phase (n = 72) and validation phase (n = 178). The possible physiological roles of the identified miRNAs were further investigated using bioinformatics analysis. Three miRNAs (miR-222-3p, miR-150-5p, and miR-423-5p) were identified as differentially expressed among groups; miR-222-3p and miR-423-5p negatively correlated with T stage, lymph node metastasis status, and clinical stage. A high diagnostic accuracy (Area under curve = 0.88) was demonstrated for discriminating OL from OSCC. Bioinformatics analysis reveals that miR-423-5p and miR-222-3p are significantly over-expressed in oral cancer tissues and involved in various cancer pathways. The three-plasma miRNA panel may be useful to monitor malignant progression from OL to OSCC and as potential biomarkers for early detection of oral cancer.
Adipocyte-specific loss of PPAR gamma attenuates cardiac hypertrophy
(2016-10-06)
Fang, Xi
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Stroud, Matthew J.
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Ouyang, Kunfu
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Fang, Li
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Zhang, Jianlin
Adipose tissue is a key endocrine organ that governs systemic homeostasis. PPAR gamma is a master regulator of adipose tissue signaling that plays an essential role in insulin sensitivity, making it an important therapeutic target. The selective PPAR gamma agonist rosiglitazone (RSG) has been used to treat diabetes. However, adverse cardiovascular effects have seriously hindered its clinical application. Experimental models have revealed that PPAR gamma activation increases cardiac hypertrophy. RSG stimulates cardiac hypertrophy and oxidative stress in cardiomyocyte-specific PPAR gamma knockout mice, implying that RSG might stimulate cardiac hypertrophy independently of cardiomyocyte PPAR gamma. However, candidate cell types responsible for RSG-induced cardiomyocyte hypertrophy remain unexplored. Utilizing cocultures of adipocytes and cardiomyocytes, we found that stimulation of PPAR gamma signaling in adipocytes increased miR-200a expression and secretion. Delivery of miR-200a in adipocyte-derived exosomes to cardiomyocytes resulted in decreased TSC1 and subsequent mTOR activation, leading to cardiomyocyte hypertrophy. Treatment with an antagomir to miR-200a blunted this hypertrophic response in cardiomyocytes. In vivo, specific ablation of PPAR gamma in adipocytes was sufficient to blunt hypertrophy induced by RSG treatment. By delineating mechanisms by which RSG elicits cardiac hypertrophy, we have identified pathways that mediate the crosstalk between adipocytes and cardiomyocytes to regulate cardiac remodeling.
alpha-ketoglutarate orchestrates macrophage activation through metabolic and epigenetic reprogramming
(2017-09-01)
Liu, Pu-Ste
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Wang, Haiping
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Li, Xiaoyun
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Chao, Tung
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Teav, Tony
Glutamine metabolism provides synergistic support for macrophage activation and elicitation of desirable immune responses; however, the underlying mechanisms regulated by glutamine metabolism to orchestrate macrophage activation remain unclear. Here we show that the production of alpha-ketoglutarate (alpha KG) via glutaminolysis is important for alternative (M2) activation of macrophages, including engagement of fatty acid oxidation (FAO) and Jmjd3-dependent epigenetic reprogramming of M2 genes. This M2-promoting mechanism is further modulated by a high alpha KG/succinate ratio, whereas a low ratio strengthens the proinflammatory phenotype in classically activated (M1) macrophages. As such, aKG contributes to endotoxin tolerance after M1 activation. This study reveals new mechanistic regulations by which glutamine metabolism tailors the immune responses of macrophages through metabolic and epigenetic reprogramming.
AMP-activated Protein Kinase Phosphorylation of Angiotensin-Converting Enzyme 2 in Endothelium Mitigates Pulmonary Hypertension
(2018-08-15)
Zhang, Jiao
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Dong, Jianjie
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Martin, Marcy
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He, Ming
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Gongol, Brendan
Rationale: Endothelial dysfunction plays an integral role in pulmonary hypertension (PH). AMPK (AMP-activated protein kinase) and ACE2 (angiotensin-converting enzyme 2) are crucial in endothelial homeostasis. The mechanism by which AMPK regulates ACE2 in the pulmonary endothelium and its protective role in PH remain elusive. Objectives: We investigated the role of AMPK phosphorylation of ACE2 Ser680 in ACE2 stability and deciphered the functional consequences of this post-translational modification of ACE2 in endothelial homeostasis and PH. Methods: Bioinformatics prediction, kinase assay, and antibody against phospho-ACE2 Ser680 (p-ACE2 S680) were used to investigate AMPK phosphorylation of ACE2 Ser680 in endothelial cells. Using CRISPR-Cas9 genomic editing, we created gain-of-function ACE2 S680D knock-in and loss-of-function ACE2 knockout (ACE22/2) mouse lines to address the involvement of p-ACE2 S680 and ACE2 in PH. The AMPK-p-ACE2 S680 axis was also validated in lung tissue from humans with idiopathic pulmonary arterial hypertension. Measurements and Main Results: Phosphorylation of ACE2 by AMPK enhanced the stability of ACE2, which increased Ang (angiotensin) 1-7 and endothelial nitric oxide synthase-derived NO bioavailability. ACE2 S680D knock-in mice were resistant to PH as compared with wild-type littermates. In contrast, ACE2-knockout mice exacerbated PH, a similar phenotype found in mice with endothelial cell-specific deletion of AMPK alpha 2. Consistently, the concentrations of phosphorylated AMPK, p-ACE2 S680, and ACE2 were decreased in human lungs with idiopathic pulmonary arterial hypertension. Conclusions: Impaired phosphorylation of ACE2 Ser680 by AMPK in pulmonary endothelium leads to a labile ACE2 and hence is associated with the pathogenesis of PH. Thus, AMPK regulation of the vasoprotective ACE2 is a potential target for PH treatment.
An enhanced computational platform for investigating the roles of regulatory RNA and for identifying functional RNA motifs
(2013-01-21)
Chang, Tzu-Hao
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Huang, Hsi-Yuan
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Hsu, Justin Bo-Kai
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Weng, Shun-Long
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Horng, Jorng-Tzong
Background: Functional RNA molecules participate in numerous biological processes, ranging from gene regulation to protein synthesis. Analysis of functional RNA motifs and elements in RNA sequences can obtain useful information for deciphering RNA regulatory mechanisms. Our previous work, RegRNA, is widely used in the identification of regulatory motifs, and this work extends it by incorporating more comprehensive and updated data sources and analytical approaches into a new platform. Methods and results: An integrated web-based system, RegRNA 2.0, has been developed for comprehensively identifying the functional RNA motifs and sites in an input RNA sequence. Numerous data sources and analytical approaches are integrated, and several types of functional RNA motifs and sites can be identified by RegRNA 2.0: (i) splicing donor/acceptor sites; (ii) splicing regulatory motifs; (iii) polyadenylation sites; (iv) ribosome binding sites; (v) rho-independent terminator; (vi) motifs in mRNA 5'-untranslated region (5'UTR) and 3'UTR; (vii) AU-rich elements; (viii) C-to-U editing sites; (ix) riboswitches; (x) RNA cis-regulatory elements; (xi) transcriptional regulatory motifs; (xii) user-defined motifs; (xiii) similar functional RNA sequences; (xiv) microRNA target sites; (xv) non-coding RNA hybridization sites; (xvi) long stems; (xvii) open reading frames; (xviii) related information of an RNA sequence. User can submit an RNA sequence and obtain the predictive results through RegRNA 2.0 web page. Conclusions: RegRNA 2.0 is an easy to use web server for identifying regulatory RNA motifs and functional sites. Through its integrated user-friendly interface, user is capable of using various analytical approaches and observing results with graphical visualization conveniently. RegRNA 2.0 is now available at http://regrna2.mbc.nctu.edu.tw.
An enhanced computational platform for investigating the roles of regulatory RNA and for identifying functional RNA motifs
(2013-01-21)
Chang, Tzu-Hao
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Huang, Hsi-Yuan
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Hsu, Justin Bo-Kai
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Weng, Shun-Long
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Horng, Jorng-Tzong
Background: Functional RNA molecules participate in numerous biological processes, ranging from gene regulation to protein synthesis. Analysis of functional RNA motifs and elements in RNA sequences can obtain useful information for deciphering RNA regulatory mechanisms. Our previous work, RegRNA, is widely used in the identification of regulatory motifs, and this work extends it by incorporating more comprehensive and updated data sources and analytical approaches into a new platform. Methods and results: An integrated web-based system, RegRNA 2.0, has been developed for comprehensively identifying the functional RNA motifs and sites in an input RNA sequence. Numerous data sources and analytical approaches are integrated, and several types of functional RNA motifs and sites can be identified by RegRNA 2.0: (i) splicing donor/acceptor sites; (ii) splicing regulatory motifs; (iii) polyadenylation sites; (iv) ribosome binding sites; (v) rho-independent terminator; (vi) motifs in mRNA 5'-untranslated region (5'UTR) and 3'UTR; (vii) AU-rich elements; (viii) C-to-U editing sites; (ix) riboswitches; (x) RNA cis-regulatory elements; (xi) transcriptional regulatory motifs; (xii) user-defined motifs; (xiii) similar functional RNA sequences; (xiv) microRNA target sites; (xv) non-coding RNA hybridization sites; (xvi) long stems; (xvii) open reading frames; (xviii) related information of an RNA sequence. User can submit an RNA sequence and obtain the predictive results through RegRNA 2.0 web page. Conclusions: RegRNA 2.0 is an easy to use web server for identifying regulatory RNA motifs and functional sites. Through its integrated user-friendly interface, user is capable of using various analytical approaches and observing results with graphical visualization conveniently. RegRNA 2.0 is now available at http://regrna2.mbc.nctu.edu.tw.
An expert system to identify co-regulated gene groups from time-lagged gene clusters using cell cycle expression data
(2010-03-15)
Wu, Li-Ching
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Huang, Jhih-Long
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Horng, Jorng-Tzong
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Huang, Hsien-Da
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生物資訊及系統生物研究所
Motivation: The analysis of time series gene expression data can provide us with the opportunity to find co-regulated genes that show a similar expression patterns under a contiguous subset of experimental conditions. However, these co-regulated genes may behave almost independently under other conditions. Furthermore, the similarity in the expression pattern might be time-shifted. In that case, we need to be concerned with grouping genes that share similar expression patterns under a contiguous subset of conditions and where the similarity in expression pattern might have time lags. in addition, to be considered a time-shifted similar pattern, because co-regulated genes in a biological process may show a periodic pattern in their cell cycle expression, we also should group genes with periodic similar patterns over multiple cell cycles. If this is carried out, we can regard such grouped genes as cell-cycle regulated genes. Results: We propose a method that follows the q-cluster concept [Ji, L., & Tan, K. L. (2005). Identifying time-lagged gene clusters using gene expression data. Bioinformatics, 21(4), 509-516] and further advances this approach towards the identification of cell-cycle regulated genes using cell cycle microarray data. We used our method to Cluster a microarray time series of yeast genes to assess the statistically biological significance of the obtained clusters we used the p-value obtained from the hypergeometric distribution. We found that several clusters provided findings suggesting a TF-target relationship. In order to test whether our method could group related genes that other methods have found difficult to group, we compared our method with other measures such as Spearman Rank Correlation and Pearson Correlation. The results of the comparison demonstrate that our method indeed could group known related genes that these measures regard as having only a weak association. (C) 2009 Elsevier Ltd. All rights reserved.
An expert system to predict protein thermostability using decision tree
(2009-07-01)
Wu, Li-Cheng
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Lee, Jian-Xin
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Huang, Hsien-Da
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Liu, Baw-Juine
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Horng, Jorng-Tzong
Protein thermostability information is closely linked to commercial production of many biomaterials. Recent developments have shown that amino acid composition, special sequence patterns and hydrogen bonds, disulfide bonds, salt bridges and so on are of considerable importance to thermostability. In this study, we present a system to integrate these various factors that predict protein thermostability. In this study, the features of proteins in the PGTdb are analyzed. We consider both structure and sequence features and correlation coefficients are incorporated into the feature selection algorithm. Machine learning algorithms are then used to develop identification systems and performances between the different algorithms are compared. In this research, two features, (E + F + M + R)/residue and charged/non-charged, are found to be critical to the thermostability of proteins. Although the sequence and structural models achieve a higher accuracy, sequence-only models provides sufficient accuracy for sequence-only thermostability prediction. (C) 2008 Elsevier Ltd. All rights reserved.
Arabidopsis Argonaute 2 Regulates Innate Immunity via miRNA393*-Mediated Silencing of a Golgi-Localized SNARE Gene, MEMB12
(2011-05-06)
Zhang, Xiaoming
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Zhao, Hongwei
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Gao, Shang
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Wang, Wei-Chi
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Katiyar-Agarwal, Surekha
Argonaute (AGO) proteins are critical components of RNA silencing pathways that bind small RNAs and mediate gene silencing at their target sites. We found that Arabidopsis AGO2 is highly induced by the bacterial pathogen Pseudomonas syringae pv. tomato (Pst). Further genetic analysis demonstrated that AGO2 functions in antibacterial immunity. One abundant species of AGO2-bound small RNA is miR393b*, which targets a Golgi-localized SNARE gene, MEMB12. Pst infection downregulates MEMB12 in a miR393b*-dependent manner. Loss of function of MEMB12, but not SYP61, another intracellular SNARE, leads to increased exocytosis of an antimicrobial pathogenesis-related protein, PR1. Overexpression of miR393b* resembles memb12 mutant in resistance responses. Thus, AGO2 functions in antibacterial immunity by binding miR393b* to modulate exocytosis of antimicrobial PR proteins via MEMB12. Since miR393 also contributes to antibacterial responses, miR393*/miR393 represent an example of a miRNA*/miRNA pair that functions in immunity through two distinct AGOs: miR393* through AGO2 and miR393 through AGO1.
Atheroprotective Flow Upregulates ITPR3 (Inositol 1,4,5-Trisphosphate Receptor 3) in Vascular Endothelium via KLF4 (Kruppel-Like Factor 4)-Mediated Histone Modifications
(2019-05-01)
He, Ming
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Huang, Tse-Shun
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Li, Shuai
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Hong, Hsiao-Chin
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Chen, Zhen
Objective-The topographical distribution of atherosclerosis in vasculature underscores the importance of shear stress inregulating endothelium. With a systems approach integrating sequencing data, the current study aims to explore the link between shear stress-regulated master transcription factor and its regulation of endothelial cell (EC) function via epigenetic modifications. Approach and Results-H3K27ac (acetylation of histone 3 lysine 27)-ChIP-seq (chromatin immunoprecipitation followed by high throughput sequencing), ATAC-seq (an assay for transposase-accessible chromatin-sequencing), and RNAseq (RNA-sequencing) were performed to investigate the genome-wide epigenetic regulations in ECs in response to atheroprotective pulsatile shear stress (PS). In silico prediction revealed that KLF4 binding motifs were enriched in the PS-enhanced H3K27ac regions. By integrating PS-and KLF4-modulated H3K27ac, we identified 18 novel PS-upregulated genes. The promoter regions of these genes showed an overlap between the KLF4-enhanced assay for transposase-accessible chromatin signals and the PS-induced H3K27ac peaks. Experiments using ECs isolated from mouse aorta, lung ECs from EC-KLF4-TG versus EC-KLF4-KO mice, and atorvastatin-treated ECs showed that ITPR3 (inositol 1,4,5-trisphosphate receptor 3) was robustly activated by KLF4 and statins. KLF4 ATAC-qPCR (quantitative polymerase chain reaction) and ChIP-qPCR further demonstrated that a specific locus in the promoter region of the ITPR3 gene was essential for KLF4 binding, H3K27ac enrichment, chromatin accessibility, RNA polymerase II recruitment, and ITPR3 transcriptional activation. Deletion of this KLF4 binding locus in ECs by using CRISPR-Cas9 resulted in blunted calcium influx, reduced expression of endothelial nitric oxide synthase, and diminished nitric oxide bioavailability. Conclusions-These results from a novel multiomics study suggest that KLF4 is crucial for PS-modulated H3K27ac that allow the transcriptional activation of ITPR3. This novel mechanism contributes to the Ca2+-dependent eNOS (endothelial nitric oxide synthase) activation and EC homeostasis.
Bacterial alterations in salivary microbiota and their association in oral cancer
(2017-11-28)
Lee, Wei-Hsiang
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Chen, Hui-Mei
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Yang, Shun-Fa
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Liang, Chao
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Peng, Chih-Yu
Oral squamous cell carcinoma (OSCC) is the most common malignant neoplasm of the oral cavity and the fourth leading malignancy and cause of cancer-related death in the male population of Taiwan. Most cases are detected at advanced stages, resulting in poor prognosis. Therefore, improved detection of early oral health disorders is indispensable. The involvement of oral bacteria in inflammation and their association with OSCC progression provide a feasible target for diagnosis. Due to the nature of oral neoplasms, the diagnosis of epithelial precursor lesions is relatively easy compared with that of other types of cancer. However, the transition from an epithelial precursor lesion to cancer is slow and requires further and continuous follow-up. In this study, we investigated microbiota differences between normal individuals, epithelial precursor lesion patients, and cancer patients with different lifestyle habits, such as betel chewing and smoking, using next-generation sequencing. Overall, the oral microbiome compositions of five genera, Bacillus, Enterococcus, Parvimonas, Peptostreptococcus, and Slackia, revealed significant differences between epithelial precursor lesion and cancer patients and correlated with their classification into two clusters. These composition changes might have the potential to constitute a biomarker to help in monitoring the oral carcinogenesis transition from epithelial precursor lesion to cancer.
Bacterial Communities in Semen from Men of Infertile Couples: Metagenomic Sequencing Reveals Relationships of Seminal Microbiota to Semen Quality
(2014-10-23)
Weng, Shun-Long
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Chiu, Chih-Min
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Lin, Feng-Mao
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Huang, Wei-Chih
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Liang, Chao
Some previous studies have identified bacteria in semen as being a potential factor in male infertility. However, only few types of bacteria were taken into consideration while using PCR-based or culturing methods. Here we present an analysis approach using next-generation sequencing technology and bioinformatics analysis to investigate the associations between bacterial communities and semen quality. Ninety-six semen samples collected were examined for bacterial communities, measuring seven clinical criteria for semen quality (semen volume, sperm concentration, motility, Kruger\'s strict morphology, antisperm antibody (IgA), Atypical, and leukocytes). Computer-assisted semen analysis (CASA) was also performed. Results showed that the most abundant genera among all samples were Lactobacillus (19.9%), Pseudomonas (9.85%), Prevotella (8.51%) and Gardnerella (4.21%). The proportion of Lactobacillus and Gardnerella was significantly higher in the normal samples, while that of Prevotella was significantly higher in the low quality samples. Unsupervised clustering analysis demonstrated that the seminal bacterial communities were clustered into three main groups: Lactobacillus, Pseudomonas, and Prevotella predominant group. Remarkably, most normal samples (80.6%) were clustered in Lactobacillus predominant group. The analysis results showed seminal bacteria community types were highly associated with semen health. Lactobacillus might not only be a potential probiotic for semen quality maintenance, but also might be helpful in countering the negative influence of Prevotella and Pseudomonas. In this study, we investigated whole seminal bacterial communities and provided the most comprehensive analysis of the association between bacterial community and semen quality. The study significantly contributes to the current understanding of the etiology of male fertility.
Biclustering of transcriptome sequencing data reveals human tissue-specific circular RNAs
(2018-01-19)
Liu, Yu-Chen
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Chiu, Yu-Jung
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Li, Jian-Rong
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Sun, Chuan-Hu
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Liu, Chun-Chi
Background: Emerging evidence has been experimentally confirmed the tissue-specific expression of circRNAs (circRNAs). Global identification of human tissue-specific circRNAs is crucial for the functionality study, which facilitates the discovery of circRNAs for potential diagnostic biomarkers. Results: In this study, circRNA back-splicing junctions were identified from 465 publicly available transcriptome sequencing samples. The number of reads aligned to these identified junctions was normalized with the read length and sequence depth for each sample. We generated 66 models representing enriched circRNAs among human tissue transcriptome through biclustering algorithm. The result provides thousands of newly identified human tissue-specific circRNAs. Conclusions: This result suggests that expression of circRNAs is not prompted by random splicing error but serving molecular functional roles. We also identified circRNAs enriched within circulating system, which, along with identified tissue-specific circRNAs, can serve as potential diagnostic biomarkers.